It is known in the prior art that various peptides having specific sequences have various physiological activities. Examples of such peptides include tripeptides Val-Pro-Pro and Ile-Pro-Pro. These tripeptides may be found in a milk fermented with lactic acid bacteria and are known to have a strong angiotensin converting enzyme (referred to hereinbelow as ACE) inhibitory activity, and show a strong anti-hypertensive activity on spontaneous hypertension rats (SHR), as well as anti-hypertensive effect on hypertensive patients (J. Dairy Sci. 1995, 78:777-783; J. Dairy Sci. 1995, 78:1253-1257; Am. J. Clin. Nutr. 1996, 64:767-771). Further, it is reported that the tripeptides Val-Pro-Pro and Ile-Pro-Pro also have an anti-stress activity (JP-A-11-100328).
As methods for producing the tripeptides Val-Pro-Pro and/or Ile-Pro-Pro, an example of efficient production with lactic acid bacteria fermentation has been reported (JP-A-11-98978). However, lactic acid bacteria fermentation ceases before complete digestion because of low pH as a result of lactic acid fermentation, which results in a large amount of undigested casein. Further, the cultured liquid produced by the lactic acid bacteria fermentation contains a large amount of lactic acid which may cause disadvantage upon processing the cultured liquid into a variety of products. For example, the co-existing lactic acid prevents formation of powders, and it is thus required to remove acids from cultured liquid for forming powders.
Another possible method for producing the tripeptides is an enzymological method. Compared to the lactic acid fermentation method, the enzymological method is expected to have advantages such as improved yield of the peptides, stable production, reduced production steps and number of operating persons, and no generation of lactic acid. However, upon digesting proteins by the enzymological method for producing the tripeptides Val-Pro-Pro and/or Ile-Pro-Pro, digestion reactivity by peptidase is considerably low at amino acid sequences containing Pro such as Xaa Pro or Pro Xaa (Xaa represents any amino acid). Therefore, it is difficult to digest the sequence Pro-Xaa in production of the tripeptides Val-Pro-Pro and/or Ile-Pro-Pro by the enzymological method, and higher yield than that in lactic acid fermentation method has not been obtained with the enzymological method. For example, JP-A-11-100328 discloses a method in which milk casein is treated with proteinase and then with carboxypeptidase, as an example of the method for obtaining the tripeptides Val-Pro-Pro and/or Ile-Pro-Pro. However there is no disclosure of the specific treating steps. JP-A-11-100328 merely discloses in the Examples other methods than the enzymological method such as lactic acid fermentation.
As the method for producing useful peptides other than the tripeptides Val-Pro-Pro and/or Ile-Pro-Pro by digestion of proteins with the combination of proteinase and peptidase, there have been proposed a method for obtaining a tripeptide Leu-Pro-Pro having ACE inhibitory activity from gamma-zein (Japanese Patent No. 2873327), and a method for obtaining a peptide Tyr-Pro-Phe-Pro-Gly-Pro-Ile-Xaa-Asn from beta-casein (JP-A-6-128287). However, these method have not been made practicable because of insufficient yield and instability for industrial production.